Fifteen to twenty percent of the adult population have serum cholesterols in excess of 240 mg/dl (moderate hypercholesterolemia) and are at increased risk of coronary heart disease. Elevated serum cholesterol usually results from an increase in low density lipoprotein (LDL), the major blood borne carrier of cholesterol. Approximately 50% of the cases of primary hypercholesterolemia may be attributable to genetic factors. The best known genetic disorder associated with elevated plasma cholesterol is familial hypercholesterolemia, which accounts for only 2% of cases of hypercholesterolemia. The candidate and sponsor have created a training program, including course work and bench research, that will prepare the candidate to investigate mutations in apolipoproteins associated with hypercholesterolemia. The sponsor has access to an inbred strain of pigs with spontaneous hypercholesterolemia and premature atherosclerosis not caused by a mutation in the LDL receptor. There is strong evidence that these animals have a mutation in apolipoprotein(apo)-B, the major protein moiety of LDL. The grant proposes to apply a novel technique to identify the causative mutation(s) in apo-B. Monoclonal antibodies to the specific domain(s) containing the mutation(s) will be raised by tolerization to normal pig LDL and immunization to mutant pig LDL. The domains will be mapped by immunoblotting of proteolytic fragments of apo-B. The corresponding portion of apo-B gene will be cloned and sequenced to determine the mutation. Clinical studies have suggested that mutations in apo-B may be a relatively common cause of hypercholesterolemia in humans. In Phase II the candidate will apply the techniques learned in studying the pigs to human subjects with moderate hypercholesterolemia. The outlined program of coursework and research will give the candidate excellent training in lipid biochemistry, cell biology and molecular genetics. The proposal will have a significant impact on our understanding of the mechanisms of hypercholesterolemia and prepare the candidate for a career of independent scientific investigation.